Summary
The objective of this study was to explore the role of 5-HT3 receptors in modulating potassium (K+)-evoked release of [3H]-acetylcholine ([3H]-ACh) from superfused slices of rat entorhinal cortex previously loaded with [3H]-choline. Rat entorhinal cortices were cross-chopped into 300 μm slices, superfused with oxygenated Krebs buffer containing 2.5 mmol/1 Ca2+ and stimulated with two consecutive exposures of 20 mmol/l K+ for 4 min (S1 and S2, respectively). Compounds were added 20 min before S2 stimulation and remained in the superfusion buffer for the duration of the experiment. The S2/S1 ratio was then calculated.
Stimulated release of [3H]-ACh was dependent on extracellular Ca2+ and K+ concentration. In Sprague Dawley rats, 2-methyl-5-HT (10-9−10-6 mol/l), in the presence of 1 μmol/l ritanserin or 1 gmmol/l ondansetron, had no influence on K+-evoked release of [3H]-ACh. In slices prepared from Hooded Lister rats, 2 μmol/l 5-HT but not 2-Me-5-HT significantly (P<0.05) inhibited K+-evoked [3H]-ACh release only 17% in the presence of 1 μmol/l ritanserin. However, 2 μmol/l 2-Me-5-HT plus 1 nmol/l ondansetron had no effect. High performance liquid chromatography coupled to electrochemical detection (HPLC-ECD) was used to monitor endogenous release of ACh in the above conditions to confirm data from the radiolabelled experiments. No significant inhibition or increase in K+-evoked ACh release was observed with either 5-HT3 receptor agonists or antagonists. 2-Me-5-HT (10−9 – 10−5 mol/l) or 1-(m-chlorophenyl)-biguanide (10−9 – 10−5 mol/l), when added simultaneously at the S2 stimulation, in the presence of 1 όl/l methysergide, also showed no effect on [3H]ACh release.
In entorhinal cortex slices from aged Wistar rats, neither 1-(m-chlorophenyl)-biguanide (2 or 10 μol/l) nor 2-Me-5-HT (2 μmol/l) in combination with ritanserin (1 μmol/l) or ondansetron (1 nmol/l) elicited any effect on K+-evoked [3H]-ACh release. However, release of [3H]-ACh was inhibited by carbachol (10 μmol/l) and adenosine (10 μmol/l). DuP 996 (3,3-bis(4- pyridinyl-methyl)-1-phenylindolin-2-one) (10−7 – 10−5 mol/l), a known releaser of ACh, markedly augmented K+-evoked [3H]-ACh release.
These studies have failed to confirm the postulated role of 5-HT3 receptors in modulating cortical ACh release in rat entorhinal cortex slices and suggest that a critical reexamination of the interaction of 5-HT3 receptor and cortical cholinergic function needs to be addressed.
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Abbreviations
- 5-HT:
-
serotonin
- ACh:
-
acetylcholine
- HPLC-ECD:
-
high performance liquid chromatography - electrical chemical detection
- EGTA:
-
ethylene glycol bis(β-aminoethyl ether)-N′,N′-tetraacetic acid
- 2-ME-5-HT:
-
2-methyl-5-hydroxytryptamine
- DuP 996:
-
(3,3-bis(4pyrindinylmethyl)-1-phenylindolin-2-one)
References
Barnes JM, Barnes NM, Costall B, Naylor RJ, Tyers MB (1989) 5-HT3 receptors mediate inhibition of acetylcholine release in cortical tissue. Nature (Lond) 338:762–763
Barnes JM, Costall B, Coughlan J, Domeney AM, Gerrard PA, Kelly M E, Naylor RJ, Onaivi ES, Tomkins DM, Tyers MB (1990) The effects of ondansetron, a 5-HT3 receptor antagonist, on cognition in rodents and primates. Pharmacol Biochem Behav 35:955–962
Beani L, Bianchi C, Siniscalchi A, Sivilotti L, Tanganelli S, Veratti E (1984) Different approaches to study acetylcholine release: endogenous ACh vs tritium efflux. Naunyn- Schmiedeberg's Arch Pharmacol 328: 119–126
Bianchi C, Siniscalchi A, Beani L (1986) The influence of 5-hydroxy-tryptamine on the release of acetylcholine from guinea-pig ex-vivo and in vitro. Neuropharmacology 25:1043–1049
Bianchi C, Siniscalchi A, Beani L (1990) 5-HT1A agonists increase and 5-HT3 agonist decrease acetylcholine efflux from the cerebral cortex of freely moving guinea pigs. Br J Pharmacol 101:448–452
Costall B, Naylor RJ, Tyers MB (1990) The psychopharmacology of 5-HT3 receptors. Pharmac Ther 47:181–202
Derkach V, Surprenant A, North RA (1989) 5-HT3 receptors are membrane ion channels. Nature 339:706–709
Domeney AM, Costall B, Gerrard PA, Jones DN, Naylor RJ, Tyers MB (1991) The effect of ondansetron on cognitive performance in the marmoset. Pharmacol Biochem Behav 38:169–175
Ferkany J, Coyle J (1983) Evoked release of aspartate and glutamate: disparities between prelabeling and direct measurement. Brain Res 278:279–282
Fozard JR (1984) Neuronal 5-HT receptors in the periphery. Neuropharmacology 23:1473–1486
Fozard JR (1987) 5-HT3 receptors and cytotoxic drug-induced vomiting. TIPS 8:44–45
Fozard JR (1992) 5-HT3 receptors in the context of the multiplicity of 5-HT receptors. In: Hamon, M (ed) Central and peripheral 5-HT3 receptors. Academic Press, London
Fozard JR, Host M (1982) Selective inhibition of the Bezold-Jarisch effects of 5-HT in the rat by antagonists of neuronal 5-HT receptors. Br J Pharmacol 77:520p
Herdon H, Strupish J, Nahorski S (1985) Differences between the release of radiolabelled and endogenous dopamine from superfused rat brain slices: effects of depolarizing stimuli, amphetamine and synthesis inhibition. Brain Res 348:309–320
Hoyer D, Schoeffter P (1991) 5-HT receptors: subtypes and second messengers. J Receptor Res 11:197–214
Jones BJ, Costall B, Domeney AM, Kelly ME, Naylor RJ, Oakley NR, Tyers MB (1988) The potential anxiolytic activity of GR38032F, a 5-HT3-receptor antagonist. Br J Pharmacol 93:985–993
Maricq AV, Petterson AS, Brake AJ, Myers RM, Julius D (1991) Primary structure and functional expression of the 5-HT3 receptor, a serotonin-gated ion channel. Science 254:432–437
Maura G, Andrioli GC, Cavazzani P, Raiteri M (1992) 5-Hydroxytryptamine3 receptors sited on cholinergic axon terminals of human cerebral cortex mediate inhibition of acetylcholine release. J Neurochem 58:2334–2337
Nickolson VJ, Tam SW, Myers MJ, Cook L (1990) DuP 996 (3,3-bis(4pyrindinylmethyl)-1-phenylindolin-2-one) enhances the stimulus-induced release of acetylcholine from rat brain in vitro and in vivo. Drug Dev Res 19:285–300
Peters JA, Lambert JJ, Malone HM (1991) Physiological and pharmacological aspects of 5-HT3 receptor function. In: Stone TW (ed) Aspects of synaptic transmission: UP, galanin, opioid, autonomic, 5-HT. Taylor and Francis, London, pp 283–313
Potter PE, Meek JL, Neff NH (1983) Acetylcholine and choline in neuronal tissue measured by HPLC with electrochemical detection. J Neurochem 41:188–194
Richardson BP, Engel G (1986) The pharmacology and function of 5-HT3 receptors. Trends Neurosci 7:424–428
Schlicker E, Betz R, Gothert M (1989) Investigation into the age-dependence of release of serotonin and noradrenaline in the rat brain cortex and of autoreceptor-mediated modulation of release. Neuropharmacology 28:811–815
Siniscalchi A, Beani L, Bianchi C (1990) Different effects of 8-OH-DPAT, a 5-HT1A receptor agonist, on cortical acetylcholine release, electrocortigram and body temperature in guinea and rats. Eur J Pharmacol 175:219–223
Smith PK, Krohn RI, Hermanson GT, Mallia AK, Gartner FH, Provenzano MD, Fujimoto EK, Goeke NM, Olson BJ, Klenk DC (1985) Measurement of protein using bicinchoninic acid. Anal Biochem 150:76–85
Takei N, Nihonmatsu I, Kawamura H (1989) Age-related decline of acetylcholine release evoked by depolarizing stimulation. Neurosci Letts 101:182–186
Zoltay G, Cooper JR (1990) Ionic basis of inhibitory presynaptic modulation in rat cortical synaptosomes. J Neurochem 55:1008–1012
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A preliminary report of this work was presented at the 1992 Federation of American Societies for Experimental Biology, April 6–9, Anaheim, California, USA (The FASEB J 6A1559)
Correspondence to R. M. Johnson at the above address
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Johnson, R.M., Inouye, G.T., Eglen, R.M. et al. 5-HT3 receptor ligands lack modulatory influence on acetycholine release in rat entorhinal cortex. Naunyn-Schmiedeberg's Arch Pharmacol 347, 241–247 (1993). https://doi.org/10.1007/BF00167441
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DOI: https://doi.org/10.1007/BF00167441